The goal of this project is to understand the roles of electrolytes in cardiac function by directly quantitating their distribution among the anatomical compartments of heart muscle. This will be accomplished by electron microprobe xray microanalysis (EPMA) of tissue sections from rabbit and rat ventricular muscle. Isolated myocytes from the same species will also be studied and compared with intact muscles. The focus is on the external and internal calcium (Ca) pools believed to be primarily involved in beattobeat excitationcontraction coupling. (1) Two Ca pools at cell surface may be related to contractility, namely, the sarcolemma and the interstitium. The latter has never been characterized but is potentially a much larger source of contractile Ca. These two regions will be characterized in situ with respect to the effects of varying Ca concentrations, cationic uncouplers of contractility, inotropic transients, and such agents as dodecyl sulphate and dimethonium. (2) The sarcoplasmic reticulum (SR) may be the major immediate source of contractile Ca, but this has yet to be directly proven in intact muscle. Ca concentration and content will be directly estimated both for the junctional SR and the entire SR. Ca release from the SR during a single heart beat will be evaluated. The effects of ryanodine on SR Ca content will be measured, and the relationship between the strength of the first postrest beat and SR Ca content will be assessed. The question is whether Ca coming directly from the SR might be adequate to explain measured force development. The uniqueness of this application is related to the use of EPMA as a tool for direct measurement of electrolytes in locations where they cannot be directly measured by any other technique. By carrying out such measurements in the subcellular structures believed to be most involved in supplying contractile Ca, it will be possible for the first time to directly evaluate hypotheses about their specific roles. More direct knowledge of how specific subcellular regions control contractility is fundamental to progress in cardiac physiology, pathology and pharmacology.